Method of treating symptoms of common cold, allergic rhinitis and infections relating to the respiratory tract

ABSTRACT

The present invention relates to methods of treating conditions and/or symptoms related to common cold of the upper and/or lower respiratory tract and/or eyes. In particular the invention relates to the methods of treating conditions and/or symptoms related to common cold comprising administration of a flavonoid or administration of a flavonoid in combination with a metal. The invention furthermore describes compositions comprising a metal and a flavonoid useful for the treatment of conditions and/or symptoms relates to common cold.

This is a continuation of Ser. No. 10/363,430 filed Sep. 22, 2003, nowU.S. Pat. No. 7,166,640, which is a 371 of PCT/DK01/00515 filed Jul. 23,2001.

The prior application(s) set forth above are hereby incorporated byreference in their entirety.

FIELD OF THE INVENTION

The present invention relates to the use of a therapeutically effectiveamount of a flavonoid and/or a flavonoid derivative such as for exampletroxerutin or Veneruton® and/or a pharmaceutically acceptable saltthereof, together with a pharmaceutically acceptable carrier for thepreparation of a medicament for treatment of one or more conditionsrelated to common cold of the upper and/or lower respiratory tractand/or eyes. Such conditions comprises “common cold”, a virus infectionor bacterial infection related to the syndrome of common cold, anallergic condition having one or more symptoms similar with the symptomsof a common cold for example allergic rhinitis initiated by rhinovirusinfection, asthma like exacerbations and/or other abnormal airwayfunctions derived from various dysfunctions of the immune system, suchas for example hay fever or the like.

The present invention further relates to a medicament for preventionand/or treatment of infections and optionally inflammations accompanyinginfections of the respiratory tract initiated by microorganisms. Theinvention relates more specifically to a medicament comprising aflavonoid and/or a flavonoid derivative, for example Venoruton® orTroxerutin, as the active substance in said medicament.

The present invention further relates to a medicament comprising zincand a flavonoid for the treatment of conditions relating to common coldand/or symptoms relating to common cold, as well as to a method oftreatment of conditions relating to common cold or symptoms relating tocommon cold involving administration of zinc and a flavonoid.

BACKGROUND ART

In the prior art no fast working and efficient composition has beenprovided for preventing and/or treating common colds initiated by viralinfections caused by the so-called cold viruses, such as rhino virus,corona virus, adenovirus, coxsackie virus, RS-virus, echovirus or othercold viruses yielding the usual well known cold syndromes in patients.Practically all humans suffer 2 to 3 times a year from infections in theupper respiratory passages, such as cold and flu. In general, in Denmarkthe majority of common colds occurring in September, October andNovember are caused by rhinovirus infection, whereas the majority ofcommon cold occurring in January, February and March are caused byCoronavirus infections.

Furthermore, there is a great need for effective remedies in theincreasing number of patients suffering from allergic syndromes, forexample asthma, which may be initiated by common cold viruses,especially the rhinovirus.

Recent observations from a polymerase chain reaction (PCR)-study(Johnston, 1993) with naturally rhinovirus infected persons indicatesthat the actual range for rhinovirus infections involved in common coldsyndrome probably is at least twofold higher, compared to findingsobtained via the traditional cell culture techniques (40%). Thisindicates that up to 70-75% of all patients suffering from common coldshave a rhinovirus infections ongoing either as a single infection orco-infection (Spector, 1995).

It has been estimated that the average pre-school child experiences 6-10upper respiratory infections or common colds per year whereas theaverage adult experiences 2-4 (Sperber, 1989). The effects of the commoncold can be uncommonly disruptive, forcing otherwise normal persons tomiss work, school, etc. Individuals who are at increased risks, such asindividuals suffering from bronchitis or asthma, may also experience alife-threatening exacerbation of their underlying conditions. Theaverage annual expenditure for various cold treatments exceeds USD 2billion in the United States, alone (Spector, 1995); in the EU a similarfigure is expected.

Currently, there is no efficient treatment to offer common coldpatients. Some offered treatments may even worsen the cold; for example,it has been demonstrated that the administration of aspirin andacetaminophen may have detrimental effects on cold treatment,neutralising antibodies and even increase nasal problems (Graham, 1990).Oral alpha-agonist may relieve congestions in many individuals andantihistamines may sometimes be helpful (Spector, 1995) but no real cureis observed Prevention or treatment with artificial soluble receptorshas not been as successful as hoped (Hayden, 1988); several trialstreating common cold patients with interferon have been completelynegative (Monto, 1989; Sperber, 1989). Plecamil® which recently (March,2000) was investigated in several trials and which inhibits the bindingof the rhinovirus via its attachment site, termed also negatively.

All these trials, which involved treatment of the syndrome common coldwere negative, despite the fact that adequate drug concentrations werepresent in the nose of the treated persons. These results indicate thatreversal of the pathogenic events in rhinovirus colds requires more thanjust the inhibition of viral replication.

Unfortunately, research in development of novel strategies to treatcommon cold is complicated by the fact human rhinoviruses only have beenreported to infect primates successfully and hence no practical animalmodel has been developed for rhinovirus infections (Rotbart, 2000).

The development of natural and experimentally induced rhinovirusinfections in normal persons are initiated by selected events which canbe considered to occur sequentially. The steps in the rhinoviruspathogenesis are believed to include viral entry into the outer nose,mucociliary transport of virus to the posterior pharynx, and initiationof infection in ciliated and non-ciliated epithelial cells of the upperairway. Viral replication peaks on average within 48 h of initiation ofinfection and persists for up to 3 weeks; Infection is followed byactivation of several inflammatory mechanisms, which may include releaseor induction of interleukins, bradykinins, prostaglandins and possiblyhistamine, including simulation of parasympathetic reflexes (thecytokines may counteract each other at certain levels resulting in avery complex pathway). The resultant clinical illness is arhinosinusitis, pharyngitis, and bronchitis, which on average lasts oneweek (Gwaltney, 1995).

Occasionally, a secondary bacterial or microbial infection may followsubsequently to the viral infection and a sustained and more seriousinflammation may result.

Previously, it was believed that the major part of the virus wasproduced in the upper nose region and excreted (Winther, 1993a).However, subsequent studies, comparing recovery of virus innasopharyngeal wash specimens, nasal swabs and pharyngeal swabs showedthat the nasopharyngeal wash specimens was consistently superior to theother two specimens in yielding virus (Cate, 1964). From a series ofin-depth investigations (Winther, 1984a; Winther, 1984b; Winther, 1984c;Turner, 1984; Farr, 1984; Hayden, 1987; Winther, 1987a; Winther, 1987b;Winther, 1993b; Arruda, 1995; Winther, 1998) it was concluded that:

-   (i) the virus was first recovered, at the highest concentrations,    from the nasopharynx before it could be recovered in the upper nose    region (turbinates).-   (ii) no evidence for rhinovirus induced damage of the surface    ciliary lining of the inferior turbinate was noted which is in    agreement with other investigators suggesting that the virus may be    transported to the nasopharynx in the overlaying mucus by    mucociliary clearence.-   (iii) there was a significant increase of the influx of neutrophils    in the same area as in (ii)-   (iv) infection of the lining of the nasal cavity was not uniform    after intranasal inoculation and seemed not to result in any cell    damage at all, cf. (ii) above.-   (v) the rate of viral shedding in the nasopharynx was high by day 1    (post infection), whereas cold symptoms did not peak until day 3.    The symptoms waned during the first week, but rhinovirus was present    during the following 3 weeks.-   (vi) The increase of neutrophils correlate with the onset of    symptoms, including sore throat. The symptoms include oedema-like    symptoms which, in turn, may trigger sneezing and coughing.

In summarising the above findings, it should be stressed that thehighest concentration of virus can be recovered from the nasopharynx,and virus usually appears on the turbinate(s) one or two days later,despite the fact that virus is innoculated via the nose (in volunteers);no visible damage of the cell lining in the upper airways was everdemonstrated. Furthermore, as “sore throat” usually developssimultaneously with the appearance of virus in the nasopharynx it can bereasoned that “signal molecules” or the like (Van Damme, 1988) will bemade by the relatively few rhinovirus cells infected and that these“cytokine-like molecules” subsequently may activate the “lymphaticring”—which is located just beneath the nasopharynx—leading to thewell-known sore throat which in turn triggers a complex pattern ofinflammatory reactions, involving an array of different interferons andcytokines the interaction of which is currently under in-depthinvestigation. Some of these factors, such as for example II-1, inducefever in patients. Bradykinines per se may be responsible for the sorethroat which is frequently associated with common cold.

The fact that interferon is known to be part of the non-specific innateimmune response against viral infections in man has lead to severalpublications as a number of groups have investigated how much interferonis produced locally during viral infections of the upper-airways. One ofthe earliest and probably most thorough, in vivo, investigations in manwas performed by Cate et al. (Cate, 1969) on volunteers (healthy adultmales from federal correctional institutions in USA): the authors wereable to demonstrate that most of the persons involved, producedinterferon (as demonstrated in nasal washings) during common colds at alevel which, at least theoretically, should have been enough to blockthe viral infection, per se. It is tempting to speculate that if nointeractions from the numerous inflammatory actions (including oedemas)had taken place, the infected persons might not had experienced atraditional cold at all.

It has been demonstrated in a recent publication, that the immune systemalso takes “active part” in the spread of the inflammatory actions sinceexperimental evidence supports the notion that rhinovirus may use someof the effector cells from the immune system as a mean for spreading theinflammatory reactions to the lower airways (Gem, 1996) via initiationof local TNF-alpha production; it is tempting to speculate that theallergic rhinitis is initiated via this mechanism as it has been foundthat the pathogenesis for asthma is linked to local TNF-alpha production(Broide et al. 1992). Several quarters have thus argued that the asthmasyndromes are rhinovirus manifestations of post-infectious eventstriggered by an array of different cytokines in connection with a“switch” between the Th1 vs. Th2 response (Gem, 1999; Winther, 1998;Grünberg, 1999).

Generally speaking, air-way infections or allergic rhinitis and/orasthma may pose a serious health problems as it can be potentiallylife-threatening for susceptible groups such as elderly people withchronic airway problems or persons suffering from a deficient immunity,such as AIDS-patients, cancer patients etc. Thus, a simple method oftreating these symptoms/syndromes (and possibly also the underlyinginfections would be of immense importance).

Viral and/or other microbial infections are known to initiate a complexinflammatory response (Ginsburg, 1988) from the patient which probablyis mediated by several groups of responder cells including theneutrophile granulocytes, which are specifically increased during acold. The latter represents approximately more than 95% of all theeffector cells; each min. about 6-9 millions neutrophiles enter theupper-airways and slowly pass down the interior surfaces encompassingthe upper airways; it may be assumed that the neutrophiles, which areable to release very aggressive enzymes and toxic substances upon properstimulation will keep the bacterial load of the upper-airways to anacceptable level; the small numbers of S. pyogenes or S. aureus found innasopharynx, which otherwise is almost sterile, may stimulate theneutrophiles via the so-called super-antigens to a certain degreethereby limiting the numbers of bacteria in said areas (dynamicequilibrium/symbiosis).

According to Ihrcke and co-workers (Ihrcke, 1993) the very early stepsin a virus infection (or any other abnormality in the cell lining) canbe related to the content and metabolism of heparan sulfate proteoglycan(the major proteoglycan associated with intact endothelial cells). Thefirst element of the model derives from the observation that heparansulfate is released from the intact endothelial lining of blood vesselsduring the very first step in an inflammatory response initiated by aviral infection. Accordingly, this loss may seriously compromise thevascular integrity and result in a local edema attracting furtherneutrophiles via the up-regulation of ICAM-1 markers on the endothelialcells increasing the inflammatory response further. Thus, in a separateexperiment, activated neutrophiles were able to release 70% of allcell-associated heparan sulfate proteoglycan within one hour via thesubsequent release of heparanase. One important function of heparansulfate is the maintenance of the endothelial cell integrity. Loss ofheparan sulfate partially abrogate the barrier properties of theendothelium and contributes to the edema and exudation of plasmaproteins that characterise inflammation.

BRIEF DISCLOSURE OF THE INVENTION

According to the present invention, it is believed that the explanationfor the therapeutic failure of the said prior art antiviral therapiesis, that the viral infections per se trigger inflammatory responseswhich can not be expected to respond at all to antiviral drugs, per se.The existence of inflammatory events in rhinovirus induced common coldsis substantiated by the finding of elevated concentrations ofinflammatory mediators, such as bradykinins, IL-8 in the nasal secretionof persons with colds (Proud, 1990; Naclerio, 1988) and the partialreduction of cold symptoms by treatment with selected antiinflammatorydrugs that have no antiviral activity (Gaffey, 1988).

Flavonoids are polyphenolic compounds isolated from a wide variety ofplants with over 4000 individual compounds known. They comprise a rangeof C₁₅ aromatic compounds and are found in virtually all land-basedgreen plants. According to one theory, upon administration of flavonoidsto an individual the flavonoid molecules are build into a part of theouter layer of the endothelial cell layer and thereby cause a reductionin microvascular hyperpermeability and hence a reduction of themigration of granulocytes through the endothelial layer. Accordingly,flavonoids can be used to inhibit oedemas and to downregulateinflammatory reactions.

WO 01/03681 describes the anti-viral effect of a number of flavonoidsfor the purpose of treatment of infections, in particular viralinfections. Although several flavonoids have been shown to comprise anantiviral effect, a number of flavonoids do not comprise any antiviraleffects in laboratory tests. Never the less the present inventiondiscloses that such non-antiviral flavonoids surprisingly are veryeffective in the treatment of common cold.

Hence, it is a first objective of the present invention to use atherapeutically effective dosage of a flavonoid and/or a flavonoidderivative selected from the group comprising hydroxyethylrutosides,troxerutin, Veneruton, genistein, taxifolin, eriodyctol, catechin,epicatechingallate, pharmaceutical acceptable salts thereof andfunctional derivatives thereof, together with a pharmaceuticallyacceptable carrier for the preparation of a medicament for treatment ofone or more conditions and/or symptoms of conditions relating to acommon cold of the upper and/or lower respiratory tract and/or eyes.

It is a second objective of the present invention to provide a method oftreatment of an individual, including a human being, comprisingadministering to said individual a therapeutically effective dosage of aflavonoid and/or a flavonoid derivative selected from the groupcomprising hydroxyethylrutosides, troxerutin, Veneruton, genistein,taxifolin, eriodyctol, catechin, epicatechin, epigallocatechin,epicatechingallate, pharmaceutical acceptable salts thereof andfunctional derivatives thereof, for prevention and/or treatment of oneor more conditions and/or symptoms of conditions relating to a commoncold of the upper and/or lower respiratory tract and/or eyes.

Preferably, said flavonoid and/or flavonoid derivative does not compriseantiviral activity when tested in vitro. It is furthermore preferredthat said flavonoid is soluble in water. More preferably, said flavonoidis a hydroxyethylrutoside, which does not comprise antiviral activitywhen tested in vitro. Yet more preferably, said flavonoid and/orflavonoid derivative is selected from the group consisting oftroxerutin, Veneruton, pharmaceutical acceptable salts thereof andfunctional derivatives thereof. Most preferably, said flavonoid and/orflavonoid derivative is selected from the group consisting of troxerutinand Veneruton®. Veneruton® is a registered trademark of NOVARTIS andcomprise a mixture of hydroxyethylrutosides, wherein around 50% istroxerutin.

In one especially preferred embodiment of the present invention theflavonoid is troxerutin of the formula:

Even though treatment of common cold patients with zinc gluconatelozengers in one trial performed by Mossad et al. lead to a reduction inthe number of days the patients were suffering from common coldsymptoms, after 4 days treatment with zinc gluconate lozengers the totalsymptom score was still more than 50% of the original symptom score.Interestingly, the present invention discloses a synergistic effectbetween administration of flavonoids and the administration of a metal,i.e. a combination therapy with flavonoids and metal is much moreefficient than either one alone.

Hence, it is a third objective of the present invention to provide apharmaceutical composition comprising a pharmaceutical effective amountof a flavonoid and/or a flavonoid derivative and/or a pharmaceuticalacceptable salt thereof, as well as a pharmaceutical acceptable metaland/or metal salt and/or metal complex.

It is a further objective of the present invention to provide a kit ofparts comprising a pharmaceutical effective amount of a flavonoid and/ora flavonoid derivative and/or a pharmaceutical acceptable salt thereof,as well as a pharmaceutical acceptable metal and/or metal salt and/ormetal complex.

It is yet a further objective of the present invention to use atherapeutically effective amount of a flavonoid and/or a flavonoidderivative and/or a pharmaceutical acceptable salt thereof, togetherwith a therapeutically effective amount of a metal and/or metal saltand/or metal complex and a pharmaceutically acceptable carrier for thepreparation of a medicament for treatment of one or more conditionsand/or symptoms relating to a common cold of the upper and/or lowerrespiratory tract and/or eyes.

It is another objective of the present invention to provide a method oftreatment of an individual, including a human being, comprisingadministering to said individual either simultaneously as separate orcombined formulation or sequentially in any order, a therapeuticeffective amount of a flavonoid and/or a flavonoid derivative and/or apharmaceutical acceptable salt thereof as well as a pharmaceuticalacceptable amount of a metal and/or metal salt and/or metal complex forameliorating, curative and/or prophylactic therapy of one or moreconditions and/or symptoms relating to a common cold of the upper and/orlower respiratory tract and/or eyes.

In combination with metal, flavonoid and flavonoid derivatives accordingto the present invention includes flavonoids of the general formula:

or the general formula:

Wherein

-   R2′ can be selected from: —H    -   —OH-   R3′ can be selected from: —H    -   —OH    -   —OCH₃    -   —OCH₂CH₂OH-   R4′ can be selected from: —H    -   —OH    -   —OCH₃    -   —OCH₂CH₂OH-   R5′ can be selected from: —H    -   —OH    -   —OCH₃    -   —OCH₂CH₂OH-   R3 including R3₁ and R3₂ can be selected from:    -   —H    -   —OH    -   —O-rutinose    -   —O-glucoside    -   —O-glucose-p-coumaric acid    -   —SOH    -   —O-rhamnose-   R4 can be selected from: —(O)    -   —OH-   R5 can be selected from: —H    -   —OH    -   —O—CH₂CH₂OH-   R6 can be selected from: —H    -   —OH    -   —OCH₃-   R7 can be selected from: —H    -   —OH    -   —O-glucose    -   —OCH₃    -   —OCH₂CH₂OH    -   —O-glucuronic acid    -   —O-rutinose    -   —O-rhamnoglucoside-   R8 can be selected from: —H    -   —OH

Furthermore, flavonoid and/or flavonoid derivatives could bestereoisomers of the above mentioned. Additionally flavonoid and/orflavonoid derivatives could be dimers comprising two flavonoid subunits.

Additionally, flavonoids and/or flavonoid derivatives of the presentinvention to be used in combination with metal could be any flavonoidand/or flavonoid derivative known to the person skilled in the art. Forexample such flavonoid and/or flavonoid derivative could be any of theflavonoid and/or flavonoid derivative mentioned in WO 01/03681, which ishereby incorporated in its entirety by reference.

Preferably, the flavonoid and/or flavonoid derivatives are selected frommolecules with the above general formulas with the proviso,

-   that when R3′ is selected from —OH    -   —OCH₃    -   —OCH₂CH₂OH-   then R5′ is selected from —H-   and when R5′ is selected from —OH    -   —OCH₃    -   —OCH₂CH₂OH-   then R3′ is selected from —H

Semi-synthetic flavonoids are also within the scope of the presentinvention.

Preferably, the flavonoid according to the present invention could beselected from the group consisting of: troxerutin, venoruton,hydroxyethylrutosides, hesperitin, naringenin, nobiletin, tangeritin,baicalein, galangin, genistein, quercetin, apigenin, kaempferol,fisetin, rutin, luteolin, chrysin, taxifolin, eriodyctol, catecithin,epicatechin, epigallocatechin, epicatechin gallate, epigallocatechingallate, flavone, sideritoflavone, hypolaetin-8-O-GI, oroxindin,3-hydroxyflavone, morin, quercetagetin-7-O-GI, tambdletin, gossypin,hipifolin, naringin, leucocyanidol, amentoflavone and derivativesthereof and mixtures thereof.

More preferably, one or more of the R chains are —OCH₂CH₂OH, yet morepreferably, at least two R chains are —OCH₂CH₂OH, most preferably threeR chains are —OCH₂CH₂OH.

Preferably, said flavonoid and/or flavonoid derivative does not compriseantiviral activity when tested in vitro. Furthermore, it is preferredthat said flavonoid is soluble in water. More preferably, said flavonoidis a hydroxyethylrutoside, which does not comprise antiviral activitywhen tested in vitro. Yet more preferably, said flavonoid and/orflavonoid derivative is selected from the group consisting oftroxerutin, Veneruton, pharmaceutical acceptable salts thereof andfunctional derivatives thereof.

In one especially preferred embodiment of the present invention theflavonoid derivative to be used is troxerutin of the formula:

Mixtures of more than one flavonoid and/or flavonoid derivative is alsocomprised within the present invention. For example such a mixture maycomprise 2, such as 3, for example 4, such as 5, for example 6, such as7, for example 8, such as 9, for example 10, such as more than 10different flavonoids. Preferably, such a mixture comprise 8 to 10different flavonoids.

In one preferred embodiment the flavonoid derivatives according to theinvention comprises a mixture of mono-, di-, tri- andtetrahydroxyethylrutosides. More preferably, the mixture comprise 1% to15% monohydroxyethylrutoside, such as from 5% to 10%monohydroxyethylrutoside, and from 25% to 50% dihydroxyethylrutoside,such as from 30% to 38% dihydroxyethylrutoside, and from 30% to 70%trihydroxyethylrutoside, such as from 45% to 55% trihydroxyethylrutosideand from 1% to −20% tetrahydroxyethylrutoside, such as from 3% to 12%tetrahydroxyethylrutoside. Most preferably, said mixture ofhydroxyethylrutosides is Venoruton.

FIGURES

FIG. 1. The number of “events” in each category (symptoms 1-4) werecounted every day. Day 0: no treatment; day 1=24 h treatment, etc.Symptom 4 (very strong), symptom 3 (strong), symptom 2 (not pleasant),symptom 1 (minimal symptoms), symptom 0 (no symptoms):

FIG. 2. The total scores from all patients under the symptom “malaise”were calculated for each day and graphed vs. a so-called “non-treated”group taken from Jackson et al, 1958 (day 2 in Jackon's study is hereused as day 0)

FIG. 3. The scores for the symptom “sore throat” were calculated foreach day as in the preceding figure.

FIG. 4. The corresponding scores for the “sneezing” symptom werecalculated as in the preceding figures as an average from 7 patients.

FIG. 5. The mean total scores from a control trial (Hertz) and the trialdescribed in example 1 (KB) are compared as %-reduction in mean totalsymptom score per patient.

FIG. 6. Average symptom score per patient during a 3 days treatment withpreparation C (ImmumaxZn).

FIG. 7. The antiviral activity of natural HulFN-α vs. Rhinovirus-T39

DETAILED DESCRIPTION OF THE INVENTION

According to the present invention conditions relating to a common coldof the upper and/or lower respiratory tract and/or eyes comprises commoncold, a viral infection and/or a bacterial infection of the upper and/orlower respiratory tract and/or eyes, rhinitis, an allergic conditionhaving one or more symptoms similar with the symptoms of a common coldfor example allergic rhinitis initiated by rhinovirus infection, asthmalike exacerbations and/or other abnormal airway functions derived fromvarious dysfunctions of the immune system, such as for example hay feveror the like.

Furthermore, conditions relating to a common cold may comprise secondarybacterial infection(s) that follow soon after a primary viral infection.Secondary bacterial infections may for example be initiated by thenormal bacterial flora present in the upper and/or lower respiratorytract and/or eyes.

Symptoms of conditions relating to common cold can be selected from thegroup comprising, but is not limited to: coughing, sneezing, musclepain, sore throat, hoarseness, irritated throat, headache, malaise,chilliness, nasal discharge, nasal obstruction, pain relating to thesinuses, fever, rhinitis, swelling of mucosal membranes, pharyngitis,astma, and acute as well as chronic bronchitis.

In the present invention the upper respiratory tract includes the mouth,nose, sinuses, throat, and the respiratory tract to epiglottis. Thelower respiratory tract includes the rest of the bronchial treeincluding the bronchioles and lung vacuoles. The invention also relatesto the treatment of eye symptoms related to the condition of therespiratory tract in that the condition may involve the mucosal liningof the respiratory tract as well of the eyes. By the term treatment asused herein is also meant prevention of symptoms whether the preventionis in fact a decrease in the development of symptoms or a prevention ofthe symptoms to arise in first place, e.g. upon exposure to infection.

According to the present invention a pharmaceutically effective amountor a therapeutically effective amount is to be understood as an amountsufficient to induce a desired biological result. The result can bealleviation of the signs, symptoms, or causes of a disease, for exampleof common cold, preferably, the result is a significant alleviation ofsigns, symptoms or causes of common cold. For example, an effectiveamount is generally that which provides either subjective relief ofsymptoms or an objectively identifiable improvement as noted by theclinician or other qualified observer, preferably such a relief ofsymptoms is a significant relief. The relief may for example beevaluated based on a symptom score as disclosed herein in the examples.Accordingly, effective amounts can vary widely depending on theindividual, on the disease or symptom to be treated.

Most common cold patients produce interferon following infection of therespiratory tract (Cate et al., 1969), which per se in principle shouldbe sufficient to alleviate the infection.

Hence, in one preferred aspect of the present invention the treatment ofa viral infection is not to be regarded as a direct antiviral effect butas a modification or inhibition of cytokines or other factors relevantfor the establishment or continuation of a viral infection located inthe mucosal membrane of the respiratory tract or eyes. Furthermore, thetreatment preferably inhibits inflammation processes in the mucosalmembrane of the respiratory tract or eyes and thereby alleviatessymptoms of common cold. Accordingly, the invention relates to use of aflavonoid and/or a flavonoid derivative for the treatment of symptoms ofviral infection of the upper and/or lower respiratory tract and/or eyes,wherein the flavonoid and/or flavonoid derivative has no antiviraleffect in vitro.

Thus, in one preferred embodiment of the present invention the flavonoidand/or flavonoid derivatives does not comprise an antiviral or antibacterial effect in vitro. In vitro antiviral and/or antibacterialeffect can be determined in various laboratory tests. Preferably, suchlaboratory tests comprise a cultured cell line capable of being infectedwith the bacteria or virus to be tested as well as said bacteria orvirus. More preferably, said cultured cell line is WISH cells and saidvirus is a rhinovirus selected from the group consisting of: rhinovirus1A, rhinovirus 15 and rhinovirus 39. Most preferably antiviral effect isdetermined using the MTS method as described in example 1. Whenantiviral effect is measured according to the MTS method as described inexample 1, a protection of less than 10%, preferably less than 7.5%,more preferably less than 5%, even more preferably less than 3%, mostpreferably less than 2% is to be regarded as no antiviral effect invitro.

Preferably, the dffect of the flavonoid and/or flavonoid derivative isclosely related to the living organism such as the effect is amodulatory effect on specific factors and biological reactions relatedto the affected mucosal membrane. The precise mechanisms are currentlynot known.

Very often common cold is initiated by, associated with or followed by aviral infection which is involved in the common cold or symptoms of thecommon cold. In one embodiment of the present invention the conditionrelating to common cold is associated with a viral infection of theupper and/or lower respiratory tract and/or eyes.

The virus infection which a common cold is most often associated with orinitiated by, is infection by one or more virus selected from the groupconsisting of: adenoviruses, parvoviruses, picornaviruses, reoviruses,orthomyxoviruses, paramyxoviruses, arenaviruses, caliciviruses,coronaviruses, orthomyxoviruses, rhinovirus, influenza virus, includinginfluenza virus type A and B, echovirus, respiratory syncytial virus(RSV), and coxsackie virus. Rhinovirus is the most common virusidentified in relation to common cold. The term rhinovirus is meant tocomprise any rhinovirus for example any of the rhinoviruses 1-113.However, very often the above virus may be present in individuals withno symptoms of common cold. Preferably, the virus infection associatedwith common cold according to the present invention is infection byrhinovirus or coronavirus.

Very often the common cold is associated with or followed by a bacterialinfection, which is involved in the common cold or symptoms of thecommon cold. Such a bacterial infection may in one embodiment of thepresent invention be a secondary infection following a primary infectionwith for example a virus. In one embodiment of the present invention thecondition relating to common cold is associated with a bacterialinfection of the upper and/or lower respiratory tract and/or eyes.

The bacterial infection which may be associated with a common cold orwith the symptoms thereof is most often infection by one or morebacteria selected from Streptococcus pheumoniae, StreptococcusHaenmolyticae, Haemophilus influenxae, and Moraxella catarrhalis.

Furthermore, common cold may be initiated by a microbial infection. Sucha microbial infection may lead to similar inflammatory responses asviral infections involving the same effector cells for exampleneutrophiles. Accordingly, such microbial infections may be treated in afashion similar to viral infections associated with common cold.

Many allergic reactions are associated with symptoms similar to thesymptoms of a common cold and it has surprisingly been shown that suchsymptoms of an allergic disorder may also be effectively treated by themethod and use as disclosed herein. Hence, in one embodiment of thepresent invention the condition relating to common cold is an allergicdisorder.

The allergic conditions according to the present invention is preferablyselected from rhinitis, asthma, acute and chronic bronchitis, and hayfewer, and the most common symptoms in this respect is one or moresymptom selected from nasal discharge, nasal congestion, sneezing,cough, swelling of mucosal membranes, rhinitis. More preferably, theallergic condition according to the present invention is selected fromthe group consisting of rhinitis and hay fewer. In a further aspect ofthe present invention the individual may have relief from the symptomsbased on a decreasing effect of said flavonoid derivatives on themucosal swelling associated with the infection or condition mentionedherein. In a still further aspect the present invention encompass acuteallergic reactions related to insect bites and sticks and in a stillfurther aspect to the allergic reactions from food or other allergensleading to swelling of the mucosa of the mouth and/or throat in suchacute reactions.

It is furthermore contained within the present invention to treatallergic conditions that is initiated by one or more agents selectedfrom the group consisting of: pollution, house dust, common dust mitesuch as Dermatophagoides Farinae or Dermatophagoides Pteronyssinus,pollen such as grass pollen, tree pollen or weed pollen, mold, animaldanders or feathers, fungal spores and chronic inhalation of forexample, wheat flour.

Accordingly, the conditions related to common cold of the presentinvention could be an infection or common cold or allergic conditioncharacterised by one or more symptoms selected from the groupcomprising: coughing, sneezing, muscle pain, sore throat, hoarseness,irritated throat, headache, malaise, chilliness, nasal discharge, nasalobstruction, pain relating to the sinuses, rhinitis, swelling of mucosalmembranes, pharyngitis, astma, and acute as well as chronic bronchitis.

When the condition relating to common cold is an allergic condition,preferably such a condition is treated by administration of flavonoidwithout simultaneous administration of metal to the individual in needthereof. More preferably said flavonoid is selected from the groupconsisting of troxerutin and Veneruton®.

The classical common cold results in symptoms, which lasts forapproximately one week. However, in certain cases conditions relating tocommon cold results in symptoms, which lasts for much longer. Such longlasting common colds for example last for more than 10 days, such asmore than 2 weeks, such as more than 3 weeks, for example more than onemonth, such as more than 6 weeks. Individual suffering from long lastingcommon cold are preferably treated by administration of flavonoidwithout simultaneous administration of metal. More preferably saidflavonoid is selected from the group consisting of troxerufin andVeneruton®.

In contrast, individuals suffering from a classical common cold whereintreatment is initiated 1 to 5 days following the onset of common coldsymptoms, preferably 1 to 3 days following the onset of common coldsymptoms are preferably treated by administration of both a flavonoidand metal according to the present invention.

In one preferred embodiment the flavonoid and/or flavonoid derivative isnot able to potentiate interferon mediated antiviral activity.Preferably, determination of potentiation of interferon mediatedantiviral activity is performed using a laboratory test measuringantiviral effect as described herein above. Such laboratory testpreferably includes measuring the antiviral effect of interferon in thepresence and absence of said flavonoid and/or derivatives. Morepreferably such test is performed as described in example 3 and 4.

The interferons could be any interferon known to the person skilled inthe art. Such interferon could be derived from a mammal including ahuman being. Such interferon could be naturally occuring interferonand/or recombinant interferon. Preferably such interferon could beselected from the group consisting of: IFN-α, IFN-β, IFN-γ and nativehuman leucocyte interferon. More preferably, such interferon could beHuIFN-α-2b.

The effective dosage of said flavonoids and/or flavonoids derivativesand/or a pharmaceutically acceptable salt thereof is preferably from 5to 5000 mg daily. More preferably, the effective dosage is from 10 mg to4000 mg, such as from 30 mg to 3000 mg, even more preferably from 40 mgto 2000 mg daily, yet more preferably, from 50 mg to 1000 mg daily.

Furthermore, the effective dosage of said flavonoids and/or flavonoidsderivatives and/or a pharmaceutically acceptable salt thereof could be adosage equivalent of a dosage of troxerutin of from 5 mg to 5000 mgdaily.

The effective dosage of Venoruton or troxerutin or a pharmaceuticallyacceptable salt or a functional derivative thereof is from 5 to 5000 mg.In general the effective dosage is from 10 mg to 4000 mg, such as from30 mg to 3000 mg, preferably from 40 mg to 2000 mg daily, morepreferably, from 50 mg to 1000 mg daily, yet more preferably from 50 to500 mg daily, most preferably from 100 to 300 mg daily.

The administration of flavonoids and/or flavonoid derivatives accordingto the present invention is preferably a very frequent administrationduring the day. Accordingly, the daily dosage may be administered individed dosages of 1 to 36 individual dosages daily preferably 2 to 24times daily, more preferably 3 to 12 times daily, such as 5 to 8 timesdaily, for example around 6 times daily. Preferably, the first 2 dosesare administrated simultaneously. The specific number of dailyapplications may be correlated to the individual way of administrationand the severity of the symptom in question. The preferred treatment isa treatment where the medicament is present in the mucosal membrane asconstant as possible due to the theory that the individual factorsinvolved in the maintenance of the symptoms are constantly produced inthe affected mucosal membrane during the illness.

In one embodiment the flavonoids or the composition or kit-of-partscomprising flavonoids and metals according to the present invention areadministrated in combination with a second treatment such as incombination with an antiviral treatment including treatment againstinfluenza such as TaMiFlu®, treatment against rhinitis such as Picovir®;or treatment with antibodies against streptococcus; or treatment withinterferons (alpha, beta or gamma) and mixtures thereof. The antiviralagents include TamiFlu or other neuraminidase inhibitors or rimantadineor antibodies against RSV.

In another embodiment of the present invention the second treatment isadministration of an anti-microbial agent. Preferably, theanti-microbial agent is distinct and specific, however theanti-microbial agent may also be a general antibiotic. In particular, ananti-microbial agent may be administrated to treat conditions associatedwith bacterial infections.

However, the flavonoids according to the present invention may beadministrated alone or in combination with a metal (see below). Inparticular, the flavonoids according to the present invention arepreferably not administrated in combination with a vitamin.

In one preferred embodiment the flavonoids are comprised in acomposition or a kit of parts that further comprises a therapeuticeffective amount of a metal and/or metal salt and/or complex orderivatives thereof.

The metal according to the present invention is preferably selected fromthe group consisting of zinc, manganese, cadmium, cobalt, iron andselenium. The metal may for example be in the form of Zn²⁺, Mn²⁺, Cd²⁺,Co²⁺, Fe²⁺ and Se²⁺. Most prefer is zinc. Preferably zinc is Zn²⁺, givenin the form of a salt and/or complex or derivatives thereof.

Within the scope of the present invention, zinc could be in any suitableform for example as ZnGluconate, as Zn(acetate)₂, as Zn²⁺ aminochelates,as Zn²⁺ amino acid chelates, as Zn²⁺ DL-methionine, as Zn²⁺L-methionine, as histidine derivatives or as a complex with amino acidsin combination with histidine, or the like such as for examplePolaPreZinc®. Furthermore zinc could be in the form of zinc sulfate,zinc chloride, Nitric-acid zinc, phosphoric-acid zinc, ulmin acid zinc,zinc fluoride, zinc iodide, a zinc hydroxide, zinc carbonate, a zincchromate, benzoic-acid zinc, zinc acetate, p-aminobenzoic-acid zinc,p-dimethylamino benzoic-acid zinc, p-zinc phenolsulfonate, p-methoxycinnamic-acid zinc, lactic-acid zinc, gluconic-acid zinc, citric-acidzinc, salicylic-acid zinc, a zinc stearate, lauric-acid zinc,myristic-acid zinc, Oleic-acid zinc, 2,5-pyridine dicarboxylic-acidzinc, 2,6-pyridine dicarboxylic-acid zinc, 4-pyridine dicarboxylic-acidzinc, 2,4-dicarboxy pyridine zinc, 3-hydroxy-2-carboxy pyridine zinc,3-n-propoxy-2-carboxy pyridine zinc, 3-n-hexyloxy-2-carboxy pyridinezinc, 5-n-propoxy-2-carboxy pyridine zinc, 5-n-butoxy-2-carboxy pyridinezinc, 5-(2-ethyl-hexyloxy)-2-carboxy pyridine zinc, 6-n-butoxy-2-carboxypyridine zinc, 3-methoxy-2-carboxy pyridine zinc, 5-methoxy-2-carboxypyridine zinc, 6-methoxy-2-carboxy pyridine zinc, 6-n-hexyloxy-2-carboxypyridine zinc, 3-methyl-2-carboxy pyridine zinc, 4-methyl-2-carboxypyridine zinc, 4-tert-butyl-2-carboxy pyridine zinc, 5-methyl-2-carboxypyridine zinc, 5-n-hexyl-2-carboxy pyridine zinc, 3-n-undecyl-2-carboxypyridine zinc, 4-n-undecyl-2-carboxy pyridine zinc, 5-n-butyl-2-carboxypyridine zinc, 6-n-undecyl-2-carboxy pyridine zinc,4-nitroglycerine-2-carboxy pyridine zinc, 5-hydroxy-2-carboxy pyridinezinc, 4-fluoro-2-carboxy pyridine zinc, 2 -carboxy pyridine N-oxidezinc, picolinic-acid zinc, Nicotinic-acid zinc, nicotinamide zinc,3,4-dihydroxy benzoic-acid zinc, Screw histidine zinc, hinokitiol zinc,protoporphyrin zinc, porphyrin zinc or picolinic-acid amide zinc.

It is contained within the present invention that zinc could be acombination of the above mentioned zinc salts and/or a zinc complexes.Such combination could comprise two or more sorts. Preferably zinc isselected from the group consisting of Zn²⁺ aminochelates, Zn²⁺ aminoacid chelates, Zn(acetate)₂, Zn²⁺ DL-methionine, Zn²⁺ L-methionine,ZnGluconate and PolaPreZinc®. Preferably, zinc is in the form ofZnGluconate or PolaPreZinc®.

The effective dosage of Zinc depends upon the form of zinc componentwhich is adminstrated. Preferably between 0.1 mg and 500 mg Zn²⁺ isadministrated, such as between 0.5 mg and 250 mg, for example between 1mg and 150 mg, such as between 5 mg and 100 mg, for example between 10mg and 50 mg per dose. If the zinc compound is ZnGluconate, preferablybetween 5 mg and 1000 mg, more preferably between 10 mg and 500 mg, evenmore preferably between 10 mg and 100 mg, yet more preferably between 20mg and 80 mg, even more preferably between 30 mg and 70 mg, mostpreferably around 50 mg ZnGluconate is administrated per dose. If thezinc compound is PolaPreZinc, preferably between 1 mg and 500 mg, morepreferably between 5 mg and 250 mg, even more preferably between 10 mgand 100 mg, most preferably around 25 mg.

The administration of a flavonoid and/or a flavonoid derivative and/or apharmaceutical acceptable salt thereof and a pharmaceutical acceptableamount of a metal and/or metal salt and/or metal complex may be eithersimultaneously as separate or combined formulations or it may besequential.

It is preferred to present flavonoids and/or flavonoid derivativesand/or metals according to the present invention in the form of apharmaceutical formulation. Accordingly, the present invention furtherprovides pharmaceutical formulations, either as a single composition oras a kit of parts, for medicinal application, which comprises aflavonoid and/or flavonoid derivative as well as a metal and/or metalsalt and/or metal complex according to the present invention or apharmaceutically acceptable salts thereof, as herein defined, and apharmaceutically acceptable carrier therefore.

The pharmaceutical formulations according to the present invention maybe prepared by conventional techniques, e.g. as described in Remington:The Science and Practice of Pharmacy 1995, edited by E. W. Martin, MackPublishing Company, 19th edition, Easton, Pa. The phamaceuticalformulation may have any form known to the person skilled in the art.For example the pharmaceutical formulation may be in the form of asolution, dispersion, emulsion, suspension, bioadhesive andnon-bioadhesive gel, powder, micropheres, tablets, lozenges, chewingtablets, chewing gum, pills, capsules, cachets, suppositories,dispersible granules, drops, sprays, aerosols, insufflators, inhalators,patches, a lollipop, ointment, lotion, cream, foam, implant, syrup orbalm. The skilled person may select the appropriate administration formbased on the common knowledge within the field of delivery systems forpharmaceuticals.

It is believed that the optimal effect is obtained by a direct topicalapplication of the flavonoids and/or metals according to the presentinvention on the mucosal membrane in question. Accordingly, it ispreferred that the administration is topical administration directly tothe mucosal membrane, more preferably, to the mucosal membrane of theupper and/or lower respiratory tract and/or of the eyes, even morepreferably the mucosal membrane of the oral cavity. The formulationshould generally be distributed to a major part of the mucosal involvedin the specific condition or symptom to be treated.

The pharmaceutical composition and/or the kit of parts according to thepresent invention usually comprise pharmaceutically acceptable carriers,which can be either solid or liquid. Carrier can be one or moresubstances which may also act as diluents, flavouring agents,solubilisers, lubricants, suspending agents, binders, preservatives,wetting agents, tablet disintegrating agents, or an encapsulatingmaterial. Such carriers include pharmaceutical grades of mannitol,lactose, starch, magnesium stearate, sodium saccharine, talcum,cellulose, glucose, lactose, pectin, dextrin, starch, gelatin, sucrose,magnesium carbonate, tragacanth, methylcellulose, sodiumcarboxymethylcellulose, a low melting wax, cocoa butter, and the like.Preferably, the pharmaceutical carrier is Magnesium stearate. Inaddition, the pharmaceutical formulations may comprise colorants,flavours, stabilisers, buffers, artificial and natural sweeteners,dispersants, thickeners, solubilising agents, and the like.

In powders, the carrier is a finely divided solid, which is a mixturewith the finely divided active components. In tablets, the activecomponents are mixed with the carrier having the necessary bindingcapacity in suitable proportions and compacted in the shape and sizedesired. The powders and tablets preferably contains from one to aboutseventy percent of the active compound.

Formulations suitable for topical administration in the mouth includelozenges comprising active agents in a flavoured base, usually sucroseand acacia or tragacanth; pastilles comprising the active ingredient inan inert base such as gelatin and glycerin or sucrose and acacia; andmouthwashes comprising the active ingredient in a suitable liquidcarrier. In one preferred embodiment the lozenges comprise sorbitoland/or peppermint oil.

The compounds of the present invention may be formulated for nasaladministration. The solutions or suspensions are applied directly to thenasal cavity by conventional means, for example with a dropper, pipetteor spray. The formulations may be provided in a single or multidoseform. In the latter case of a dropper or pipette this may be achieved bythe patient administering an appropriate, predetermined volume of thesolution or suspension. In the case of a spray this may be achieved forexample by means of a metering atomizing spray pump.

The compounds of the present invention may be formulated for aerosoladministration, particularly to the respiratory tract and includingintranasal administration. The compound will generally have a smallparticle size for example of the order of 5 microns or less. Such aparticle size may be obtained by means known in the art, for example bymicronization. The active ingredient is provided in a pressurized packwith a suitable propellant such as a chlorofluorocarbon (CFC) forexample dichlorodifluoromethane, trichlorofluoromethane, ordichlorotetrafluoroethane, carbon dioxide or other suitable gas. Theaerosol may conveniently also contain a surfactant such as lecithin. Thedose of drug may be controlled by a metered valve. Alternatively theactive ingredients may be provided in a form of a dry powder, forexample a powder mix of the compound in a suitable powder base such aslactose, starch, starch derivatives such as hydroxypropylmethylcellulose and polyvinylpyrrolidine (PVP). The powder carrier will form agel in the nasal cavity. The powder composition may be presented in unitdose form for example in capsules or cartridges of e.g., gelatin orblister packs from which the powder may be administered by means of aninhaler.

Surprisingly, the present invention discloses that even though commoncold is usually caused by an infection of the upper and/or lowerrespiratory tract, it can be treated effectively by topicaladministration directly to the mucosal membrane of the oral cavity.Since administration directly to the mucosal membrane of the oral cavityis very convenient for the individual to be treated, it is aconsiderable advantage of the present

invention that administration can be performed directly to said mucosalmembrane. In addition, the present invention discloses that allergicrhinitis also can be treated by applying the compounds according to thepresent invention directly to the musocal membrane of the oral cavity.Accordingly, the compounds according to the present invention arepreferably formulated as lozenges, chewing tablets, chewing gum, drops,sprays and aerosols, which can be applied directly to the mucosalmembrane of the oral cavity. Most preferably, the compounds according tothe present invention are formulated as lozenges, which can be directlyapplied to the mucosal membrane of the oral cavity.

The individual in need of a treatment according to the invention couldbe any individual, however preferably, such individual is a human being.The individual will generally have a score relating to symptoms based onthe score system as disclosed in Patients diary, (see examples) of atleast 4 to 5, such as at least 6, preferably, at least 10, morepreferably the patient would have a score of at least 15, whereas anindividual with a score of 3 or less is not to be regarded as sick.Generally speaking a score around 5 to 6 or lower will allow the personto continue his/her work.

In a further aspect of the invention, the treatment results in adecrease in the severity of symptoms corresponding to a decrease ofscore as measured according to patients diary herein of at least 15%within 24 hours, such as least 25%, more preferably of at least 30% in24 hours from the start of the treatment. After 48 hours of treatmentthe scores is preferably decreased with at least 20% in 48 hours, suchas with at least 30%, for example with around 40% to 60%, morepreferably with at least 40%, yet more preferably with at least 50%,even more preferably with at least 60% in 48 hours from the start of thetreatment 72 hours of treatment preferably results in a decrease ofscore as measured according to Patients Diary herein of at least 30%,preferably at least 40%, more preferably at least 50%, even morepreferably at least 55%, yet more preferably at least 59%, even morepreferably at least 65%, most preferably at least 70% in 72 hours fromthe start of the treatment. However, the preferred decrease in symptomscore is dependent on the condition relating to common cold to betreated, the scheme of treatment and the individual patient.

Flavonoids are known to possess anti-oxidative properties, and accordingto one further aspect, the flavonoid is a flavonoid having a singletOxygen Quenching measured as the rate constant of ¹O₂ quenching K offrom 10⁴ to 10⁹ M⁻¹ s⁻¹. Preferably, the rate is 10⁴to 10⁸ M⁻¹ s⁻¹. Thesinglet oxygen quenching can be measured using a variety of solventsknown to the person skilled in the art. Preferably, the solvent isselected from the group consisting of CD₃OD, a mixture of CCl₄ and CH₃OHof 1:3 and CH₃CN.

EXAMPLES

Preparation A (Immumax)

Venoruton ® 50 mg Sorbitol 934 mg Peppermint oil 8 mg Magnesiumstearat10 mg In total 1000 mg

Preparation B

Troxerutin 50 mg Sorbitol 934 mg Peppermint oil 8 mg Magnesiumstearat 10mg In total 1000 mg

Preparation C (ImmuMaxZn)

Venoruton ® (Novartis) 50 mg Zn Gluconate (Fertin) 50 mg Sorbitol 882 mgPeppermint oil 8 mg Magnesiumstearat 10 mg In total 1000 mg

Patients Diary

This scheme should preferably be filled out in the evening. How are yourcurrent condition with regard to the symptoms below. In the schemebelow, please state the strength of your symptoms today by inserting anX at the appropriate place: every symptom should have points: 0 meansthat you have not had any symptoms at all; 4 means that you have had theworst symptoms available; etc. 0=no symptoms, 1=a minimum of symptoms;2=unpleasant symptoms; 3=considerably unpleasant symptoms; 4=veryunpleasant symptoms

Symptom points 0 1 2 3 4 Cough Headache Hoarseness Nasal dischargeSneezing Nasal obstruction Sore throat Irritated throat Malaise Soremuscles FeverHave you had any side effects of the treatment? Yes □ No □Specify _(——————)Do you also take any other medical treatment or other kinds of treatmentapart from this test treatment? Yes □ No □Specify _(——————)

The scheme above may preferable be used for identifying persons in needof a treatment according to the invention and to compare the effect withother treatments or placebo. A total score of 3 to 5 or less is regardedto be a normal condition.

Example 1

Virus Titrations

Rhinovirus 1A, rhinovirus 15 and rhinovirus 39 were titrated accordingto the tetra zolium salt (MTS)-method (Berg et al., 1989; Berg and Owen,2001a, Hansen et al., 1989). WISH cells were seeded in a micro tray at3000 cells per well and incubated at 37° C., 5% CO₂ overnight; thefollowing morning the medium Was replaced with 10-fold dilutions ofeither rhinovirus 1A, rhinovirus 15 or rhinovirus 39, respectively, infresh medium and the trays were incubated 4-5 days at 33° C.; amicroscopical examination confirmed that the CytoPathogenic Effect (CPE)was fully developed (CPE equal to 100%). The minimal amount of virus(i.e.: the highest dilution of the virus in question) which produced100% destruction was used as “challenge virus” in the subsequentexperiments. To quantitate the CPE in terms of % destruction, MTS (Bergand Owen, 2001a) was added to all cultures and after 3 h incubation at37° C. (without CO₂) the trays were read in a scanner as previuoslydescribed (Berg et al, 1989, Hansen et al., 1989). Control cellcultures, that were not infected with virus, were included in theexperiment; the latter gave the highest OD as these cells were notdamaged; depending on the concentration of virus added to the differentwells, the OD₄₉₂ varied, accordingly: 100% CPE yielded a low OD(<0.200);0% CPE corresponding to no infection at all (controls cell) gave a highOD(>1.200).

Example 2

No Antiviral Activity of ZnGluconate as Measured via the MTS-system.

WISH cells were seeded in wells in a microtray and incubated for 24 h at34° C., 5% CO₂; the medium was replaced with fresh medium comprising2-fold dilutions of ZnGluconate (diluted 1:10 from a 1% stock dilutions)and incubated further for 3-4 days at 33° C., 5% CO₂; on the followingday challenge virus was added and after 3-5 days at 33° C., 5% , CO₂,MTS was added and the microtray was measured in an OD-scanner (Berg etal., 1989; Hansen et al., 1989). Alternatively, instead of ZnGluconateWISH cells were incubated with other zinc salts/complexes or with theflavonoid derivatives, Troxerutin, Veneruton® or Quercetin.

No substantial protection against rhinovirus by addition of ZnGluconatecould be detected (<2% protection). The OD signals from the wells, thatwere incubated in the presence of ZnGluconate were very close to thevirus control curves (FIG. 7). Similar results were observed whentesting the antiviral effect of other zinc salts/complexes. When theflavonoid derivatives, Troxerutin and Veneruton were added to the WISHcells, they also did not show any antiviral activity (<1% protection).However Quercetin, had a moderate antiviral activity at levels not toxicto the cells (10 to 15% protection).

Example 3

Antiviral Activity of Interferon-α (rHuIFN-α-2b) Against Rhino Virus(1A, 15 or 39).

3.000 WISH cells were seeded in a microtray and on the followingmorning, the medium was replaced with 2-fold dilutions (from a 0-30units/ml stock solution) of HuIFN-α-2b (Intron A) in fresh mediumcomprising 2% serum. After incubation overnight, the medium was replacedwith fresh medium comprising challenge virus and incubated at 33° C., 5%CO₂ for 3-5 days and processed further as described in Example 2.

The results in FIG. 7 clearly demonstrates that rhinoviruses arereasonable sensitive to HuIFN-α-2b (50% protection). However, 90-100%protection can be achieved at approx. 8-15 units/mil.

Example 4

Troxerutin and Interferon-α (rHuIFN-α-2b) vs. Rhino Virus (1A, 15 or 39)

An experiment similar to Example 3 was carried out in which various, butconstant concentrations of either Troxerutin or Veneruton or otherrelevant derivatives were added together with the 2-fold dilutions ofinterferon to WISH cells. A typical example from this series ofexperiments yielded no potentiation of the interferon system, (<1%) atall at Troxerutin levels below 2.5 mg/ml. Troxerutin concentrationshigher than 2.5 mg/ml are toxic to the cells.

Example 5

Rhinitis and Common Cold

Preliminary Trial Involving 2 Persons.

Case 1

A) A 60 year old healthy man acquired a rhinitis with occasionallycoughs. The person applied one tablet (preparatin B) under his tongueand allowed the tablet to melt slowly during a period of 4-5 min. afterthe period of which the rhinitis sensation gradually disappeared within10 min. (no more rhinitis, no more coughing).

The rhinitis started again after approx. 20-30 min. and the personapplied 1-2 more tablets (preparation B) as described above after whichno more rhinitis was observed. A mild headache disappeared. No sideeffects were noticed. The rapid effect of the treatment stronglysuggests that the effect is caused by local action of the pharmaceuticalrather than by systemic action.

The same person had previously on a few occasions treated himself with1-2 lozenges (preparation B) when a mild rhinitis appeared and each timeone lozenge stopped the rhinitis.

Case 2

B) A 57 year healthy woman began coughing and an early rhinitis asdescribed above appeared; the person applied one tablet (preparation B)under her tongue and kept it for 3-5 min. When asked about the result afew hours later she reported that the rhinitis had disappeared during30-45 min. after one tablet, only. No side effects were noticed.

Preliminary Conclusion on Case 1 and 2

It appears that the oral treatments comprising 1-3 preparation B tabletshave been efficient as the early rhinitis and coughing disappeared. Thepersons did not catch new rhinitis or colds during the following 4-8weeks period (ending Dec. 15, 1999).

C) Preliminary trial involving 7 common cold patients at Doctor's Office

Based on the above results it was decided to go into a small controlledtrial involving 10-15 patients who had acquired the infection naturallyand were treated subsequently at a doctor's Office (HA). The optimalseason for the classical common cold is from September to February butunfortunately it was not possible to arrange for a small controlled“trial” before the end of March, 2000.

Patient Treatment.

Patients reporting to Doctor's Office (HA) complaining of a cold withrhinitis or a strong pharyngitis or other common cold syndromes (cf.Patient's Diary) were asked to fill out the Patients Diary (see above)which included relevant medical examinations (age, sex, patients onmedical treatment were excluded, the date for the onset of the coldsymptoms as listed in the diary—cf. enclosed copy. No specific effortsother than the usual symptoms of common cold were employed forcharacterisation of the patients enrolled. Thus most of the patients hadexperienced the cold for at least 1-2 days before reporting to theDoctor's Office. It is fair to assume that the patients probablyrepresent a group of patient with a more severe cold than others notreporting to the Office.

A total of 7 patients with the usual characteristics for common cold—cf.the enclosed copy of the Patient's Diary above—were treated with theImmuMax lozenges (preparation A) described above (50 mg Venoruton®) fora total period of 3 days, only. Each patient was instructed to fill outthe patients diary every day (day 0=1^(st) visit to the Doctor's Office)and to follow the mode of administration: one lozenge should be appliedon or under the patient's tongue and it should melt in a minimum periodof 4-5 min. (no fluid or food should be taken the next 15-20 min.) Ifnecessary, the patient could take the next lozenge after 30 min.; atotal number of 5 lozenges per day was equal to the maximal dose perday.

The patients were asked to fill out the diaries and to return them toDoctor's Office a few days after the treatment.

Results

The results of this very limited and preliminary trial are shown intables 1-8 and summarised in FIGS. 1-6. Events are defined as the numberof marks noted in the 5 different symptoms categories. The data in table1/FIG. 1, which shows the number of events per day from the 7 patientsin 5 symptom categories, supports the “early” preliminary “trial” asmost of the severe symptoms have disappeared after 48 h; after further24 h the patients have recovered almost completely. The curve forsymptom 4 (very strong) drops within the first 24 h and stays at almost0 for the remainder 48 h. The curve for symptom 3 (strong) had 15 countson day 0; the curve drops in parallel to symptom 4 curve and at day 2(=48 h treatment) no more counts with symptom 3 are reported from the 7patients. The curve for symptom 2 drops in parallel with the twopreceding curves, at day 3, three counts for symptom 2 were reported.The curve for symptom 1 reaches a plateau on day 1 and drops at day 3 to8 counts. The symptom 0 curve shows 8 counts at day 1; then it rises inparallel to symptom curve 3 during the first 24 h, but it continues torise to 45 on day 3.

In conclusion it can be stated that the fact that the symptom eventscore curve rises to a maximum for symptom zero supports the earlierfindings with the two persons who were also cured within two days time.

As each patient fills out the diary each symptom receives a score gradedfrom 0-4. These scores were compared with a “non-treated” control grouptaken from Jackson et al, Arch. Internal. Med. 101:267-278, 1958 (day 2in Jackon's study is here used as day 0); The total scores from allpatients under the syndrome “malaise” were compared with the so-called“non-treated” group The treated patients curve drops almost as astraight line in contrast to the non-treated, but infectious patientscurve. At day 3 the treated group has significantly less scores comparedto the non-treated group (FIG. 2, table 2 and 3). Similarly the scoresfor sore throat are significantly reduced after 3 days treatmentcompared to the non-treated group (FIG. 3, table 4 and 5). The scoresfor sneezing are close to zero after 3 days of treatment in contrast tothe non-treated group (FIG. 4, table 6 and 7).

Furthermore this study were compared with a controlled trial, designatedHertz, with 23 common cold patients who received a spray with anothertest drug for 6 days—the outcome was judged as negative although someminor effects were seen. This trial served as a control group fornon-treated patients (placebo). When the mean total scores from thesetwo groups are compared as percent reduction the present trial yielded a50% reduction in symptom scores each day compared to the Hertz trialshowing varying reductions. Based on this comparison it can be arguedthat the present trial yields a significant reduction in symptom scorescompared to the placebo study.

TABLE 1 x A B C D E values symptom 0 symptom 1 symptom 2 symptom 3symptom 4 day Y Y Y Y Y sum 1 0 8 6 20 15 6 55 2 1 9 22 15 8 0 54 3 2 2623 5 0 1 55 4 3 45 7 3 0 0 55 The number of events in each category(symptom 1-4) were counted every day, symptom 4 (very strong), symptom 3(strong), symptom 2 (not pleasant), symptom 1 (minimal symptoms),symptom 0 (no symptom

TABLE 2 A treated patients X values day X Y1 Y2 Y3 Y4 Y5 Y6 Y7 1 0 2 3 32 1 4 2 2 1 2 2 1 1 1 3 1 3 2 1 1 1 0 1 1 1 4 3 1 0 0 0 0 1 1 Malaisescores from patient Y1-Y7 from day 0-3

TABLE 3 B non-treated patients Y1 Y2 Y3 Y4 Y5 Y6 Y7 1 2.240 2 2.660 32.450 4 1.750 Malaise scores from control group

TABLE 4 A treated patients X values day X Y1 Y2 Y3 Y4 Y5 Y6 Y7 1 0 0 2 23 3 1 3 2 1 0 2 1 1 0 0 2 3 2 0 1 1 0 0 0 3 4 3 0 0 0 0 0 0 2 Sorethroat scores from patient Y1-Y7 from day 0-3

TABLE 5 B non-treated patients Y1 Y2 Y3 Y4 Y5 Y6 Y7 1 4.0 2 4.0 3 2.8 42.1 Sore throat scores from untreated group

TABLE 6 A treated patients X values day X Y1 Y2 Y3 Y4 Y5 Y6 Y7 1 0 1 3 23 4 0 2 2 1 0 2 1 1 2 0 2 3 2 0 1 1 0 0 0 2 4 3 0 0 0 0 0 0 1 Sneezingscores from patient Y1-Y7 from day 0-3

TABLE 7 B non-treated patients Y1 Y2 Y3 Y4 Y5 Y6 Y7 1 2.170 2 1.680 31.400 4 1.300 Sneezing scores from control group

TABLE 8 X values A % B reduction C Data D Data E Data day reduction HESet-C Set-D Set-E X K Y Y Y Y Y 1 0 0 0 2 1 50 22 3 2 50 24 4 3 48 37Percent reduction in total symptom score. K is this trial, HE is theHertz trial

Conclusions from the Preliminary Trial with 7 Patients

Based on the findings in FIGS. 1-5 it appears that the common coldpatients are cured significantly faster than the non-treated group asthe common cold period is cut down with approx. 50% from 6-7 days to 2-3days. Furthermore, the more annoying symptoms seem to be cleared alreadyduring the first 24 h.

Example 6

Prelimininary trial with 3 patients suffering from allergic airwayconditions

Case 1.

A 56-year old patient (BK/230244) who was allergic to grass and birchpollen was seeking advice at Doctor's Office; the patients had arhinitis and irritated eyes; the condition had been unchanged during thelast 2 weeks (no treatment was received during this period). The patientreceived the ImmuMax-treatment as described in example 1 and was askedto report after 5 days and to fill out the questionnaire in thepatient's diary. The total symptom score had been reduced >95% duringthe first 2-3 days of treatment.

Case 2

A 38-year old patient (CH/120962) who was notoriously allergic to birchpollen went to her Doctor's Office seek help; the exacerbations includedrhinitis and irritated eyes; the symptoms had lasted for 2-3 days; notreatment was initiated at the time the patient went to Doctor's Office.The patient received the same treatment as in case 1 and was asked tofill in patients diary and reported back 5 days later: The initialpatient score was very high for hoarseness, sore throat and scratchythroat, coughs, etc. After 2 days treatment with ImmuMaX a significantimprovement was noted as the symptom score decreased >75%.

Case 3

A 66-year old patient (ES/270334) had developed common cold symptomswhich had sustained for 2 weeks; she received ImmuMaX as described aboveand reported back 5 days later: at day 0, the symptom score was approx.50% of the total possible score; the symptom score was reduced >95%after 2-3 days of treatment.

Preliminary Conclusion.

Based on the results from this very limited group of allergic patientsit seems as if Venoruton® has a certain effect vs. allergy (pollenallergy and the like); furthermore, allergy which may have been inducedvia a slow upper airway infection (viral or bacterial or both) may alsolend itself to an efficient treatment using the ImmuMax.

Commentary

The fact treatment was given exclusively by the oral/local route alsodeserves further exploitations as this manner of administration israther surprising giving the fact that most if not all treatments vs.allergic conditions are given either as spray or systemically. Our newadministration form support the notion that one of the very early eventsinitiating allergic reactions/asthma like syndromes could be therecruitment of the neutrophile granulocytes which as a matter of fact isin constant contact with the areas affected by infectious processesand/or inflammatory responses which in turn lend support to our currenthypothesis namely, that it may be possible to control these events viadrugs as Troxerutin or the like.

Example 7

Preliminary Trial Involving 7 Common Cold Patients at Doctors OfficeTreated with ImmuMaxZn

7 patients reporting at the Doctor's office, who had experienced theusual symptoms of common cold (sore throat, coughing, sneezing, runningnose etc.) for 1-3 days, were treated. Each patient received 20ImmuMaxZn tablets (containing 50 mg veneruton and 50 mg ZnGluconate, seepreparation C) orally in the course of a 3 day treatment. On the firstday patients received 5-7 tablets. Patients were asked to keep apatient's diary (see herein above). 7 out of 7 patients responded to thetreatment with a 40-50% reduction in symptom score based on the patientdiaries.

The results are shown in FIG. 6 and table 9. The curve demonstrates amarked decrease in symptom score after treatment. The reduction insymptom score made it possible for the patients to attend their normalduties.

Example 8

Trial Including 42 Patients Suffering from Common Cold.

The trial was performed in Denmark, only adults participated in thetrial. 42 patients suffering from common cold were treated with eitherImmumax (preparation A) or ImmumaxZn (preparation C). Each patientreceived a maximum of 5 lozenges daily and a total of 20 lozenges duringthe course of treatment. Each lozenge was administrated by placing itunder/on the tongue and allowing it to dissolve slowly (3-5 min.) andthe followings 10 min. the patient did not drink or eat.

Each patient was asked to fill in the patients diary (see herein above).The results of the trial is summarised in table 10 to 17.

There is a significant decrease in symptom score after 3 days oftreatment with either Immumax or ImmumaxZn. However, when comparing theresults of table 11 and table 15 it becomes clear that patientssuffering from a typical common cold syndrome who started treatmentwithin 3 days after the on set of the symptoms respond better totreatment with ImmuMaxZn than to treatment with ImmuMax. A meanreduction in symptom score of 59% after 3 days treatment with ImmuMax isobserved, but 78% mean reduction in symptom score after 3 days treatmentwith ImmuMaxZn is observed.

Furthermore, it appears that the success of treatment is not dependenton the month in which the treatment was undertaken. Since the majorityof common colds in Denmark occuring in September, October and Novemberare caused by rhinovirus and the majority of common colds occuring inDenmark in January, February and March are caused by coronaviruses, thisindicates that the treatment is effective against different kinds ofvirus infections. Accordingly, it is likely that the effect of ImmuMaxor ImmuMaxZn is not a direct antiviral effect in vivo.

TABLE 10 Patients with a typical common cold syndrome presented within24 h treated with ImmuMaxZn Cured Pt. Days of Initial SS on (C Aller-Patient's Month No. Symptoms SS Day 3 Or NC) gen Remark Nov 21-00 <1 142 C No March 12-01 <1 6 0 C No Good effect

TABLE 11 Patients with a typical common cold syndrome presented within 3days treated with ImmuMaxZn Pt. Days Initial SS Cured Patient's MonthNo. of Symptoms SS on Day 3 (C or NC) Allergen Remark Nov. 22-00 3 11 4C No Nov. 27-00 3 14 0 C No Very good Nov. 37-00 1 10 0 C No good Dec.41-00 3 30 3 C No Dec. 42-00 1 10 7 Partial No Good ! Jan. 03-01 2 24 6Partial No Good effect ! March 07-01 2 18 6 Partial No

TABLE 12 Patients with common cold syndrome/allergy treated withImmuMaxZn Days Initial Cured Patient's Month Pt. No. of Symptoms SS SSon Day 3 (C or NC) Allergen Remark Nov. 29-00 3 15 3 C Pollen AverageNov. 31-00 14 10 5 (C) Birch Nov. 35-00 8 20 14 NC Dec. 46-00 3 10 2 CGrass Good Dec. 47-00 3 20 15 NC Grass March 08-01 2 16 5 (C) PollenGood

TABLE 13 Patients (without any allergic background) with a typicalcommon cold syndrome presented after 3 days treated with ImmuMaxZn DaysInitial SS Cured Patient's Month Pt. No. of Symptoms SS on Day 3 (C orNC) Allergen Remark Nov. 28-00 6 21 9 PR No Nov. 33-00 30 5 5 NC No Nov.34-00 4–5 29 29 NC No Nov. 36-00 64 22 23 NC No Jan. 48-00 6 9 2 C NoMarch 06-01 7 11 2 C No Good effect March 09-01 14 7 3 C No Good effect

TABLE 14 Patients with a typical common cold syndrome presented within24 h treated with ImmuMax Days Initial SS Cured Patient's Month Pt. No.of Symptoms SS on Day 3 (C or NC) Allergen Remark Oct. 17-00 <1 18 21 NCNo Nov. 30-01 <1 5 5 NC No

TABLE 15 Patients with a typical common cold syndrome presented within 3days treated with ImmuMax Days Initial SS Cured Patient's Month Pt. No.of Symptoms SS on Day 3 (C) Or NC Allergen Remark Jan. 13-00 2 20 1 C NoOct. 14-00 2 33 2 C No Nov. 15-00 2 11 9 NC No Dec. 19-00 2 32 30 NC NoNov. 20-00 2 23 9 Partial No Nov. 38-00 3 9 3 C No good March 07-01 2 186 Partial No

TABLE 16 Patients with common cold syndrome/allergy treated with ImmuMaxDays Initial SS Cured Patient's Month Pt. No. of Symptoms SS on Day 3(C) Or NC Allergen Remark Nov. 02-00 14 27 8 (C) Nov. 03-00 14 5 0 CJan. 52-00 6 15 12 NC grass Jan. 53-00 6 10 7 NC grass week

TABLE 17 Patients (without any allergic background) with a typicalcommon cold syndrome presented after 3 days treated with ImmuMax DaysInitial SS Cured Patient's Month Pt. No. of Symptoms SS on Day 3 (C) OrNC Allergen Remark Jan. 18-00 4 33 2 C no good March 25-00 14 15 1 C noDec. 39-00 8 15 3 C no average Dec. 40-00 6 17 5 C no average Dec. 43-0030 9 7 NC no Jan. 18-00 4 33 2 C no good March 25-00 14 15 1 C no

Table 10 to 17. SS symptom score, C cured, NC not cured, Pt. No. patientnumber, month indicates the month in which the treatment was performed,Days, of symptom indicates the number of days the patient was sufferingfrom common cold or a common cold related condition prior to the onsetof treatment. Day 3 is day 3 of treatment.

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1. A pharmaceutical composition, effective in vivo as an antiviralagent, comprising (1) a pharmaceutically effective amount of at leastone hydroxyethylrutoside or pharmaceutically acceptable salt thereof,said hydroxyetheylrutosides or salts thereof including troxerutin or apharmaceutically acceptable salt thereof, said amount being antivirallyeffective in vivo, and (2) an amount of zinc gluconate effective in vivoto cause the in vivo antiviral effect of the composition to exceed thatof the hydroxyethylrutoside(s), or salts thereof, of the composition,wherein said composition is suitable for topical administration to themucosal membrane of the oral cavity, and is not a solution, unless suchsolution is a spray.
 2. The pharmaceutical composition of claim 1 whichcomprises a mixture of mono-, di-, tri-and tetrahydroxyethylrutosides.3. The pharmaceutical composition of claim 1 wherein the composition isformulated as a lozenge, chewing tablet, chewing gum, spray or aerosol.4. The composition of claim 1, wherein the composition is formulated asa lozenge.
 5. The composition of claim 1 wherein the onlyhydroxyethylrutoside present in the composition is troxerutin.
 6. Thecomposition of claim 1 which is a mixture of 5-10%monohydroxyethylrutoside, 30-38% dihydroxyethylrutoside, 45-55%trihydroxyethylrutoside, and 3-12% tetrahydroxyethylrutoside.
 7. Thecomposition of claim 1 wherein the hydroxyethylrutoside provides lessthan 10% protection of WISH cells against rhinovirus when tested invitro according to the tetrazolium salt (MTS) method.
 8. Thepharmaceutical composition according to claim 1, wherein the compositionis formulated as a lozenge, chewing tablet, chewing gum, drop, spray oraerosal.
 9. A method of treatment of a condition or symptoms of acondition, wherein the condition is selected from the group consistingof common cold, viral infection of the upper and/or lower respiratorytract and/or eyes, rhinitis and hay fever, said method comprisingadministering to an individual in need thereof a therapeuticallyeffective amount of the composition according to claim
 1. 10. The methodaccording to claim 9, wherein the symptoms are one or more symptomsselected from the group consisting of cough, sneezing, muscle pain, sorethroat, hoarseness, headache, malaise, chilliness, nasal discharge,nasal obstruction, pain relating to the sinuses, rhinitus, swelling ofmucosal membranes and pharyngitis.
 11. The method according to claim 9,wherein the virus infection is caused by or associated with one or moreviruses selected from the group consisting of adenoviruses,picornaviruses, reoviruses, orthomyxoviruses, paramyxoviruses,caliciviruses, coronaviruses, orthomyxoviruses, rhinoviruses, echovirusand coxsackie virus.
 12. The method according to claim 9, wherein thevirus infection is caused by or associated with one or more virusesselected from the group consisting of coronaviruses and rhinoviruses.13. The method according to claim 9, wherein the virus infection isassociated with a secondary bacterial infection.
 14. The methodaccording to claim 13, wherein the bacterial infection is caused by oneor more bacteria selected from the group consisting of Streptococcuspheumoniae, Streptococcus Haemolyticae, Haemophilus influenxae andMoraxella catarrhalis.